Therapeutics
Abstract
Deficits in IL-2 signaling can precipitate autoimmunity by altering the function and survival of FoxP3+ regulatory T cells (Tregs) while high concentrations of IL-2 fuel inflammatory responses. Recently, we showed that the non-beta IL-2 SYNTHORIN molecule SAR444336 (SAR’336) can bypass the induction of autoimmune and inflammatory responses by increasing its reliance on IL-2 receptor α chain subunit (CD25) to provide a bona fide IL-2 signal selectively to Tregs, making it an attractive approach for the control of autoimmunity. In this report, we further demonstrate that SAR’336 can support non-beta IL-2 signaling in murine Tregs and limit NK and CD8+ T cells’ proliferation and function. Using a murine model of spontaneous type 1 diabetes, we showed that the administration of SAR’336 slows the development of disease in mice by decreasing the degree of insulitis through the expansion of antigen-specific Tregs over Th1 cells in pancreatic islets. Specifically, SAR’336 promoted the differentiation of IL-33–responsive (ST2+), IL-10–producing GATA3+ Tregs over other Treg subsets in the pancreas, demonstrating the ability of this molecule to further orchestrate Treg adaptation. These results offer insight into the capacity of SAR’336 to generate highly specialized, tissue-localized Tregs that promote restoration of homeostasis during ongoing autoimmune disease.
Authors
Fernando Alvarez, Nicole V. Acuff, Glenn M. La Muraglia II, Nazila Sabri, Marcos E. Milla, Jill M. Mooney, Matthew F. Mackey, Mark Peakman, Ciriaco A. Piccirillo
Abstract
KRAS mutations are frequent in various human cancers. The development of selective inhibitors targeting KRAS mutations has opened a new era for targeted therapy. However, intrinsic and acquired resistance to these inhibitors remains a major challenge. Here, we found that cancer cells resistant to KRAS G12C inhibitors also display cross-resistance to other targeted therapies, such as inhibitors of RTKs or SHP2. Transcriptomic analyses revealed that the Hippo-YAP/TAZ pathway is activated in intrinsically resistant and acquired-resistance cells. Constitutive activation of YAP/TAZ conferred resistance to KRAS G12C inhibitors, while knockdown of YAP/TAZ or TEADs sensitized resistant cells to these inhibitors. This scenario was also observed in KRAS G12D–mutant cancer cells. Mechanistically, YAP/TAZ protects cells from KRAS inhibitor–induced apoptosis by downregulating the expression of proapoptotic genes such as BMF, BCL2L11, and PUMA, and YAP/TAZ reverses KRAS inhibitor–induced proliferation retardation by activating the SLC7A5/mTORC1 axis. We further demonstrated that dasatinib and MYF-03-176 notably enhance the efficacy of KRAS inhibitors by reducing SRC kinase activity and TEAD activity. Overall, targeting the Hippo-YAP/TAZ pathway has the potential to overcome resistance to KRAS inhibitors.
Authors
Wang Yang, Ming Zhang, Tian-Xing Zhang, Jia-Hui Liu, Man-Wei Hao, Xu Yan, Haicheng Gao, Qun-Ying Lei, Jiuwei Cui, Xin Zhou
Abstract
Glucocorticoid-induced osteoporosis (GIOP) lacks fully effective treatments. This study investigated the role of Piezo1, a mechanosensitive ion channel component 1, in GIOP. We found reduced Piezo1 expression in cortical bone osteocytes from patients with GIOP and a GIOP mouse model. Yoda1, a Piezo1 agonist, enhanced the mechanical stress response and bone mass and strength, which were diminished by dexamethasone (DEX) administration in GIOP mice. RNA-seq revealed that Yoda1 elevated Piezo1 expression by activating the key transcription factor Hes1, followed by enhanced CaM kinase II and Akt phosphorylation in osteocytes. This improved the lacuno-canalicular network and reduced sclerostin production and the receptor activator of NF-κB/osteoprotegerin ratio, which were mitigated by DEX. Comparative analysis of mouse models and human GIOP cortical bone revealed downregulation of mechanostimulated osteogenic factors, such as osteocrin, and cartilage differentiation markers in osteoprogenitor cells. In human periosteum-derived cells, DEX suppressed differentiation into osteoblasts, but Yoda1 rescued this effect. Our findings suggest that reduced Piezo1 expression and activity in osteocytes and periosteal cells contribute to GIOP, and Yoda1 may offer a novel therapeutic approach by restoring mechanosensitivity.
Authors
Nagahiro Ochiai, Yuki Etani, Takaaki Noguchi, Taihei Miura, Takuya Kurihara, Yuji Fukuda, Hidetoshi Hamada, Keisuke Uemura, Kazuma Takashima, Masashi Tamaki, Teruya Ishibashi, Shohei Ito, Satoshi Yamakawa, Takashi Kanamoto, Seiji Okada, Ken Nakata, Kosuke Ebina
Abstract
The opioid system plays crucial roles in modulating social behaviors in both humans and animals. However, the pharmacological profiles of opioids regarding social behavior and their therapeutic potential remain unclear. Multiple pharmacological, behavioral, and immunohistological c-Fos mapping approaches were used to characterize the effects of μ-opioid receptor agonists on social behavior and investigate the mechanisms in naive mice and autism spectrum disorder–like (ASD-like) mouse models, such as prenatally valproic acid–treated mice and Fmr1-KO mice. Here, we report that low-dose morphine, a μ-opioid receptor agonist, promoted social behavior by selectively activating neurons in prosocial brain regions, including the nucleus accumbens, but not those in the dorsomedial periaqueductal gray (dmPAG), which are only activated by analgesic high-dose morphine. Critically, intra-dmPAG morphine injection counteracted the prosocial effect of low-dose morphine, suggesting that dmPAG neural activation suppresses social behavior. Moreover, buprenorphine, a μ-opioid receptor partial agonist with less abuse liability and a well-established safety profile, ameliorated social behavior deficits in two mouse models recapitulating ASD symptoms by selectively activating prosocial brain regions without dmPAG neural activation. Our findings highlight the therapeutic potential of brain region–specific neural activation induced by low-dose opioids for social behavior deficits in ASD.
Authors
Soichiro Ohnami, Megumi Naito, Haruki Kawase, Momoko Higuchi, Shigeru Hasebe, Keiko Takasu, Ryo Kanemaru, Yuki Azuma, Rei Yokoyama, Takahiro Kochi, Eiji Imado, Takeru Tahara, Yaichiro Kotake, Satoshi Asano, Naoya Oishi, Kazuhiro Takuma, Hitoshi Hashimoto, Koichi Ogawa, Atsushi Nakamura, Hidekuni Yamakawa, Yukio Ago
Abstract
The application of chimeric antigen receptor (CAR) T cell therapy in solid tumors is hindered by life-threatening toxicities resulting from on-target, off-tumor killing of nonmalignant cells that express low levels of the target antigen. Mesothelin (MSLN) has been identified as a target antigen for CAR T cell treatment of mesothelioma, lung, ovarian, and other cancers because of its high expression on tumor cells and limited expression on mesothelial cells. However, fatal off-tumor toxicity of high-affinity MSLN-targeting CAR T cells has been reported in multiple clinical trials. In this study, we constructed CARs using mutant variants of a single-domain nanobody that bind both human and mouse MSLN with a wide range of affinities and examined tumor responses and their toxicities from on-target, off-tumor interactions in mouse models. CAR T cells with low nanomolar affinity (equilibrium dissociation constant, KD) exhibited profound systemic expansion with no apparent infiltration into the tumor. With a gradual reduction of CAR affinity toward the micromolar KD, the expansion of CAR T cells became more restricted to tumors. Our preclinical studies demonstrated that high-affinity MSLN CARs were associated with fatal on-target, off-tumor toxicity and that affinity-tuned CARs rendered T cells more selective for MSLN-high tumors.
Authors
Yanping Yang, Yogindra Vedvyas, Yago Alcaina, Sydney J. Trumper, Diella S. Babu, Irene M. Min, Jacqueline M. Tremblay, Charles B. Shoemaker, Moonsoo M. Jin
Insulin mitigates acute ischemia induced atrial fibrillation and sinoatrial node dysfunction ex vivo
Abstract
Acute atrial ischemia is a well-known cause of postoperative atrial fibrillation (POAF). However, mechanisms through which ischemia contributes to the development of POAF are not well understood. In this study, ex vivo Langendorff perfusion was used to induce acute ischemia and reperfusion in the heart in order to mimic POAF. Inducibility of atrial fibrillation (AF) was evaluated using programmed electrical stimulation and confirmed with open-atrium optical mapping. Compared to the control group without ischemia, 25 minutes of ischemia substantially increased the incidence of AF. The right atrium was more susceptible to AF than the left atrium. Administering insulin for 30 minutes before ischemia and during reperfusion with 25 minutes of ischemia greatly reduced the vulnerability to AF. However, insulin treatment during reperfusion only did not show substantial benefits against AF. Optical mapping studies showed that insulin mitigates ischemia-induced abnormal electrophysiology, including shortened action potential duration and effective refractory period, slowed conduction velocity, increased conduction heterogeneity, and altered calcium transients. In conclusion, insulin reduced the risk of acute ischemia/reperfusion-induced AF via improving the electrophysiology and calcium handling of atrial cardiomyocytes, which provides a potential therapy for POAF.
Authors
Huiliang Qiu, Fan Li, Hannah Prachyl, Alejandra Patino-Guerrero, Michael Rubart, Wuqiang Zhu
Abstract
Type 1 diabetes (T1D) is characterized by the autoimmune destruction of insulin-producing beta cells and involves an interplay between beta cells and cells of the innate and adaptive immune systems. We investigated the therapeutic potential of targeting 12-lipoxygenase (12-LOX), an enzyme implicated in inflammatory pathways in beta cells and macrophages, using a mouse model in which the endogenous mouse Alox15 gene is replaced by the human ALOX12 gene. Our finding demonstrated that VLX-1005, a potent 12-LOX inhibitor, effectively delayed the onset of autoimmune diabetes in human gene replacement non-obese diabetic mice. By spatial proteomics analysis, VLX-1005 treatment resulted in marked reductions in infiltrating T and B cells and macrophages with accompanying increases in immune checkpoint molecule PD-L1, suggesting a shift towards an immune-suppressive microenvironment. RNA sequencing analysis of isolated islets and polarized proinflammatory macrophages revealed significant alteration of cytokine-responsive pathways and a reduction in interferon response after VLX-1005 treatment. Our studies demonstrated that the ALOX12 human replacement gene mouse provides a platform for the preclinical evaluation of LOX inhibitors and supports VLX-1005 as an inhibitor of human 12-LOX that engages the enzymatic target and alters the inflammatory phenotypes of islets and macrophages to promote the delay of autoimmune diabetes.
Authors
Titli Nargis, Charanya Muralidharan, Jacob R. Enriquez, Jiayi E. Wang, Kerim B. Kaylan, Advaita Chakraborty, Sarida Pratuangtham, Kayla Figatner, Jennifer B. Nelson, Sarah C. May, Jerry L. Nadler, Matthew B. Boxer, David J. Maloney, Sarah A. Tersey, Raghavendra G. Mirmira
Abstract
Radiation-induced lung injury (RILI) initiates radiation pneumonitis and progresses to fibrosis as the main side effect experienced by patients with lung cancer treated with radiotherapy. There is no effective drug for RILI. Sustained vascular activation is a major contributor to the establishment of chronic disease. Here, using a whole thoracic irradiation (WTI) mouse model, we investigated the mechanisms and effectiveness of thrombopoietin mimetic (TPOm) for preventing RILI. We demonstrated that administering TPOm 24 hours before irradiation decreased histologic lung injury score, apoptosis, vascular permeability, expression of proinflammatory cytokines, and neutrophil infiltration in the lungs of mice 2 weeks after WTI. We described the expression of c-MPL, a TPO receptor, in mouse primary pulmonary microvascular endothelial cells, showing that TPOm reduced endothelial cell–neutrophil adhesion by inhibiting ICAM-1 expression. Seven months after WTI, TPOm-treated lung exhibited less collagen deposition and expression of MMP-9, TIMP-1, IL-6, TGF-β, and p21. Moreover, TPOm improved lung vascular structure, lung density, and respiration rate, leading to a prolonged survival time after WTI. Single-cell RNA sequencing analysis of lungs 2 weeks after WTI revealed that TPOm shifted populations of capillary endothelial cells toward a less activated and more homeostatic phenotype. Taken together, TPOm is protective for RILI by inhibiting endothelial cell activation.
Authors
Jeb English, Sriya Dhanikonda, Kathryn E. Tanaka, Wade Koba, Gary Eichenbaum, Weng-Lang Yang, Chandan Guha
Abstract
Effective, reproducible, and safe delivery of therapeutics into the inner ear is required for the prevention and treatment of hearing loss. A commonly used delivery method is via the posterior semicircular canal (PSCC); however, its specific targeting within the cochlea remains unclear, impacting precision and reproducibility. To assess safety and target specificity, we conducted in vivo recordings of the pharmacological manipulations delivered through the PSCC. Measurements of auditory brainstem response (ABR), vibrometry, and vestibular behavioral and sensory-evoked potential (VsEP) revealed preserved hearing and vestibular functions after artificial perilymph injections. Injection of curare, a mechanoelectrical transducer (MET) channel blocker that affects hearing when in the endolymph, had no effect on ABR or VsEP thresholds. Conversely, injection of CNQX, an AMPA receptor blocker, or lidocaine, a Na+ channel blocker, which affects hearing when in the perilymph, significantly increased both thresholds, indicating that PSCC injections selectively target the perilymphatic space. In vivo tracking of gold nanoparticles confirmed their exclusive distribution in the perilymph during PSCC injection, supporting the pharmacological finding. Together, PSCC injection is a safe method for inner ear delivery, specifically targeting the perilymphatic space. Our findings will allow for precise delivery of therapeutics within the inner ear for therapeutic and research purposes.
Authors
Jinkyung Kim, Jesus Maldonado, Dorothy W. Pan, Patricia M. Quiñones, Samantha Zenteno, John S. Oghalai, Anthony J. Ricci
Abstract
Retinitis pigmentosa (RP) is a complex group of inherited retinal diseases characterized by progressive death of photoreceptor cells and eventual blindness. Pde6a, which encodes a cGMP-specific phosphodiesterase, is a crucial pathogenic gene for autosomal recessive RP (RP43); there is no effective therapy for this form of RP. The compact CRISPR/SaCas9 system, which can be packaged into a single adeno-associated virus, holds promise for simplifying effective gene therapy. Here, we demonstrated that all-in-one AAV-SaCas9-mediated Nrl gene inactivation can efficiently prevent retinal degeneration in a RP mouse model with Pde6anmf363/nmf363 mutation. We screened single guide RNAs (sgRNAs) capable of efficiently editing mouse Nrl gene in N2a cells and then achieved effective gene editing by using a single AAV to co-deliver SaCas9 and an optimal Nrl-sg2 into the mouse retina. Excitingly, in vivo inactivation of Nrl improved photoreceptor cell survival and rescued retinal function in treated Pde6a deficient mice. Thus, we showed that a practical, gene-independent method, AAV-SaCas9-mediated Nrl inactivation, holds promise for future therapeutic applications in patients with RP.
Authors
Zhiquan Liu, Siyu Chen, Chien-Hui Lo, Qing Wang, Yang Sun
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